Darrell Fleischman (Advisor)
Master of Science (MS)
Photosynthetic rhizobia have been studied for about 15 years now. They are now considered to be metabolically aligned with a relatively recently discovered group of bacteria, the anoxygenic aerobic phototrophs (AAP’s).Rhizobia form symbiotic relationships with plants from the Fabaceae family. Photosynthetic rhizobia not only nodulate the roots, as most other rhizobia do, but they also form nodules on the stems of certain leguminous plants. The plant provides carbon to the bacteria and the bacteria provides the plant with soluble nitrogen fixed from the biologically inert but abundant atmospheric N2. A key question regarding photosynthetic rhizobia and other AAP’s derives from the observation that photosynthesis in these organisms shuts down under anaerobic conditions. It has been proposed, and is the hypothesis of this thesis that the primary electron acceptor (QA) in the photosynthetic reaction center has a higher midpoint potential than in reaction centers found in the AAP’s counterparts, the anaerobic purple bacteria. If QA had a higher midpoint potential, it would be more labile to overreduction under anoxic conditions, and if QA is reduced, then photosynthetic electron transport is blocked. A redox titration was done to measure the midpoint potential of Q in the reaction centers of BTAi 1. This was done by observing the level of P (primary electron donor) bleaching upon excitation with bright light at different ambient redox potentials. The level of P bleaching is proportional to the fraction of QA that is not reduced, since P cannot bleach and donate an electron if QA is already reduced. Reaction centers from BTAi 1 were purified using two techniques, both involving ion exchange chromatography and one involving ammonium sulfate precipitation. Reaction centers were characterized by spectrophotometric studies, mass spectroscopy studies (MALDI TOF) and the cofactor composition was determined.Themidpoint potential of QA in BTAi 1 is –44 mV vs. SHE. The molecular weights of the subunits are very comparable to other photosynthetic reaction centers, from both aerobic and anaerobic bacteria. The pigment stoichiometry of reaction centers from BTAi1 is 2:1 bacteriochlorophyll:bacteriopheophytin. Both absorbance and light minus dark absorbance spectra are nearly identical to that found in anaerobic photosynthetic bacteria.Photosynthetic reaction centers in BTAi 1 are very similar to reaction centers of anaerobic photosynthetic bacteria. The midpoint potential of QA cannot account for its overreduction under anaerobic conditions. It is likely that AAP’s lack a key enzyme that would participate in redox homeostasis of the photosynthetic electron transport chain.
Department or Program
Department of Biochemistry and Molecular Biology
Year Degree Awarded
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